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Review
Streptococcus pneumoniae colonisation
Streptococcus pneumoniae colonisation: the key
to pneumococcal disease
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D Bogaert, R de Groot, and P W M Hermans
Streptococcus pneumoniae is an important pathogen
causing invasive diseases such as sepsis, meningitis, and
pneumonia. The burden of disease is highest in the youngest
and oldest sections of the population in both more and less
developed countries. The treatment of pneumococcal
infections is complicated by the worldwide emergence in
pneumococci of resistance to penicillin and other antibiotics.
Pneumococcal disease is preceded by asymptomatic
colonisation, which is especially high in children. The current
seven-valent conjugate vaccine is highly effective against
invasive disease caused by the vaccine-type strains.
However, vaccine coverage is limited, and replacement by
non-vaccine serotypes resulting in disease is a serious
threat for the near future. Therefore, the search for new
vaccine candidates that elicit protection against a broader
range of pneumococcal strains is important. Several
surface-associated protein vaccines are currently under
investigation. Another important issue is whether the aim
should be to prevent pneumococcal disease by eradication
of nasopharyngeal colonisation, or to prevent bacterial
invasion leaving colonisation relatively unaffected and hence
preventing the occurrence of replacement colonisation and
disease. To illustrate the importance of pneumococcal
colonisation in relation to pneumococcal disease and
prevention of disease, we discuss the mechanism and
epidemiology of colonisation, the complexity of relations
within and between species, and the consequences of
the different preventive strategies for pneumococcal
colonisation.
Lancet Infect Dis 2004; 4: 144–54
Streptococcus pneumoniae is a common cause of invasive
disease and respiratory-tract infections in more and less
developed countries. Risk groups for diseases caused by
pneumococci, such as meningitis, sepsis, and pneumonia,
include young children, elderly people, and patients with
immunodeficiencies.1 Each year, 1 million children younger
than 5 years old die from pneumonia and invasive diseases.
In the USA, the annual number of fatal pneumococcal
infections is 40 000.2 Community-acquired pneumococcal
meningitis has a very high case-fatality rate (20% and 50%
in more and less developed countries, respectively).
Depending on age, 30–60% of survivors develop long-term
sequelae including hearing loss, neurological deficits, and
neuropsychological impairment.3
Protection against pneumococcal infections is mediated
by opsonin-dependent phagocytosis. Antibody-initiated
144
complement-dependent opsonisation, which activates the
classic complement pathway, is thought to be the major
immune mechanism protecting the host against
pneumococcal infections.4 The mechanism of clearance
depends on the interaction of type-specific antibodies (IgA,
IgM, IgG), complement, and neutrophils or phagocytic cells
from lung, liver, and spleen. Functional or anatomical
asplenia and cirrhosis of the liver both predispose to severe
pneumococcal infection. Congenital deficiencies in
immunoglobulin or complement are also associated with
predisposition to pneumococcal infection.5 S pneumoniae is
part of the commensal flora of the upper respiratory tract.
Together with Moraxella cattarrhalis, Haemophilus
influenzae, Neisseria meningitidis, Staphylococcus aureus, and
various haemolytic streptococci, they colonise the
nasopharyngeal niche. Though colonisation with
pneumococci is mostly symptomless, it can progress to
respiratory or even systemic disease (figure 1). An important
feature is that pneumococcal disease will not occur without
preceding nasopharyngeal colonisation with the
homologous strain.6,7 In addition, pneumococcal carriage is
believed to be an important source of horizontal spread of
this pathogen within the community. Crowding, as occurs
in hospitals, day-care centres, and prisons, increases
horizontal spread of pneumococcal strains.8–16 Because the
highest frequency of pneumococcal colonisation and the
highest crowding index are found in young children, this
risk group is thought to be the most important vector for
horizontal dissemination of pneumococcal strains within
the community.17 Therefore, part of the strategy to prevent
pneumococcal disease focuses on prevention of
nasopharyngeal colonisation, especially in children.
Owing to the key role of nasopharyngeal colonisation in
pneumococcal disease and pneumococcal spread, we focus
in this review on the different features of nasopharyngeal
colonisation in children. To elucidate the route of
pneumococcal disease, we discuss current knowledge on the
mechanism of colonisation, the epidemiology and
determinants of pneumococcal carriage, and the status of
prevention of colonisation by means of vaccination.
DB is a paediatric resident, RdG is a paediatric infectious diseases
and immunology specialist, and PWMH is head of the Laboratory of
Paediatrics at Erasmus MC-Sophia, Rotterdam, Netherlands.
Correspondence: Dr P W M Hermans, Laboratory of Paediatrics,
Room Ee 1500, Erasmus MC-Sophia Rotterdam, Dr Molewaterplein
50, 3015 GE Rotterdam, Netherlands. Tel +31 10 408 8224;
fax +31 10 408 9486; email [email protected]
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Streptococcus pneumoniae colonisation
Airborne droplets
Nasopharyngeal
carriage
Aspiration
Local spread
Alveoli
Pneumonia
Pleura
Otitis media
Pericardium
Empyema
Blood
Empyema
Peritoneum
Peritonitis
Sinusitis
Septicaemia
Joints
Meninges
Arthritis/osteomyelitis
Meningitis
Figure 1. Pathogenic route for S pneumoniae infection. Redrawn from reference 2. Organs infected through the airborne and haematogenic routes are
depicted in blue and red, respectively.
Dynamics of nasopharyngeal colonisation
The upper respiratory tract is the ecological niche for many
bacterial species. In children, the nasopharyngeal flora
become established during the first months of life.7,18 A broad
variety of microorganisms including S pneumoniae,
H influenzae, and M catarrhalis can colonise the
nasopharyngeal niche. Every individual is likely to be
colonised with these pathogens at least once during life. In
general, there is simply asymptomatic carriage; but in some
cases, colonisation is followed by disease.19,20 Colonisation is
commonly followed by horizontal dissemination of the
pathogens to individuals in the direct environment, leading
to spread within the community.21–23 The reported rates of
bacterial acquisition and carriage depend on age,
geographical area, genetic background, and socioeconomic
conditions.11,23–26 The local host immune response has an
important regulatory role in the trafficking of pathogens in
the upper respiratory tract.27 A poor mucosal immune
response might lead to persistent and recurrent colonisation
and consequently infection, whereas a brisk local immune
response to the pathogen will eliminate colonisation and
prevent recolonisation.28,29 In general, mucosal immunity
matures earlier than systemic immunity, and is present from
the age of 6 months.28 IgG and secretory IgA antibodies
directed against capsular polysaccharides and surfaceassociated proteins have been observed in saliva of children
in response to colonisation with S pneumoniae.30,31
Nasopharyngeal colonisation is a dynamic process in
terms of the turnover of colonising species and serotypes.
Moreover, interspecies competition is thought to occur and to
THE LANCET Infectious Diseases Vol 4 March 2004
interfere with the composition of the nasopharyngeal
flora. First, the balance between the resident flora
and transient invaders is important. The resident flora,
including ␣-haemolytic streptococci, inhibit colonisation by
S
pneumoniae,
H
influenzae,
S
aureus,
and
M catarrhalis.22,28,32,33 The importance of this inhibitory role was
shown by Ghaffar and colleagues,28 who found a competitive
balance between ␣-haemolytic streptococci and S pneumoniae
and H influenzae, which could be altered by antibiotics.
A negative association between viridans streptococci and
S pneumoniae, H influenzae, and M catarrhalis has also been
reported, with the last three becoming predominant during
upper-respiratory-tract infections.22,34
Furthermore, the different pathogenic species show a
competitive relationship. In-vitro studies by Pericone and
colleagues35 showed a positive relation between
N meningitidis and S pneumoniae. Growth of S pneumoniae
increased in the presence of meningococci, a process
probably mediated by meningococcal catalase. However,
meningococcal growth was decreased in the presence of
pneumococci or pneumococcal culture supernatant. The
researchers attributed the latter effect to the presence of
pneumococcal peroxide.35 This inhibitory effect of
S pneumoniae was also observed in co-cultures with
H influenzae and M catarrhalis. Moreover, S pneumoniae can
interfere with the growth of S aureus; this effect has also been
attributed to pneumococcal hydrogen peroxide.36,37 We
showed in a cross-sectional carriage study of 3200 children
that the competition between S aureus and S pneumoniae
contributes substantially to the age-related dynamics of
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145
Review
Streptococcus pneumoniae colonisation
Table 1. Pneumococcal colonisation and serotype-distribution studies
Ref
Year
Country
Number of
children
Age
Risk group
Type of culture
Carriage (%) Coverage with 7-valent
conjugate vaccine (%)
63
1998–99
India
64
1997–99
Greece
464
2–6 months
Healthy
Transnasal
64–70*
2448
2–23 months
Healthy
Transnasal
34
65
1994–95
India
65
100
6–18 months
Healthy
Transnasal
40
46‡
66
1994–95
Finland
329
2–24 months
Healthy
67
1997
Indonesia
484
0–25 months
Healthy
Transnasal
13–43*
53
Transnasal
48
23
1999
Netherlands
535
68
1990
Kenya
69
2905
Taiwan
70
1997
USA
71
1996
Vietnam
38
2002
Netherlands
7
1995
USA
72
1988–92
Costa Rica
73
(2002)
Israel
1000
1–24 months
Healthy
Throat
2
..
74
1998–00
Italy
55
6–84 months
Healthy
Throat
24
..
75
2000
Italy
2799
0–7 years
Healthy
Throat
9
63
11
1996
Italy
1723
1–7 years
Healthy
Throat
4
..
25
2000
Turkey
1382
0–10 years
Healthy
Throat
8
60
1998–99
Switzerland
2769
0–16 years
RTI
Transnasal
66
1994–95
Finland
329
2–24 months
URTI
68
1990
Kenya
26
0–2 years
URTI
76
1992–94
Thailand
1783
0–5 years
URTI
Transnasal
35
..
66
1994–95
Finland
329
2–24 months
AOM
Transnasal
45–56*
68
77
1994–96
Israel
120
3–36 months
AOM
Transnasal
63
61
78
1998–02
Netherlands
383
1–7 years
Recurrent AOM Transnasal
55
55
79
1996
France
0–24 months
Orphanage
58
85
80
1996
Romania
1–38 months
Orphanage
Transnasal
50
98‡
68
1990
Kenya
26
0–2 years
HIV
Transnasal
20
59§
68
1990
Kenya
26
0–2 years
HIV and URTI
Transnasal
86
59§
80
1996
Romania
40
3–9 years
HIV
Transnasal
30
98‡*
70
1997
USA
85
0–14 years
HIV
Transnasal
20
..
61
1994–95
USA
312
0–18 years
SCD
Transnasal
21–11*
56
26
2905
50†
3–36 months
Healthy
Transnasal
37
56
0–2 years
Healthy
Transnasal
22
59§
0–7 years
Healthy
Transnasal
21
..
85
0–14 years
Healthy
Transnasal
19
..
911
1–16 years
Healthy
Transnasal
44
70‡
3200
1–19 years
Healthy
Transnasal
50–8*
42
306
6 months
Healthy
Not stated
23
440
1–12 months
Healthy
Not stated
71
162
3–19*
..
..
..
48–39*
49–65
Transnasal
22–45*
68
Transnasal
29
59§
Transnasal
62
1994–95
USA
278
1–19 years
SCD
Transnasal/throat
32–5*
79
23
1999
Holland
535
3–36 months
DCC
Transnasal
58
59
81
1998–99
Asia
4963
0–5 years
DCC/OPD
Transnasal
11–43
65‡
82
1999–00
Hong Kong
1978
2–6 years
DCC
Transnasal
39
..
83
1999
Italy
610
2–65 months
DCC
Not stated
15
57
72
1988–92
Costa Rica
280
2–5 years
DCC
Not stated
39
..
74
1998–00
Italy
85
6–84 months
Recurrent AOM Throat
29
..
75
1998–00
Italy
113
6–84 months
COME
Throat
35
..
84
1994
Japan
2–12 years
COME
Throat
23
..
43
RTI=respiratory-tract infection; URTI=upper-respiratory-tract infection; AOM=acute otitis media; SCD=sickle-cell disease; DCC=day-care centre; OPD=outpatient department;
COME=chronic otitis media with effusion. *Increasing with age. †Coverage for 9-valent conjugate vaccine. ‡Including cross-reactive serotypes. §Average for all isolates of the
study.
nasopharyngeal colonisation in children.38 Our findings have
been confirmed by Regev-Yochay and co-workers.39 We also
found that parallel to the age-related decline in
pneumococcal colonisation, caused by the maturation of the
immune system, there was a simultaneous increase in
S aureus carriage rate, from 10% in the first years of life to a
maximum of 50% at the age of 10 years. In addition to these
146
ecological interactions, the composition of the
nasopharyngeal niche is influenced by environmental factors
such as crowding and smoking.38 There is limited evidence
on the competition between the different pneumococcal
serotypes. For example, Lipsitch and colleagues40 used a
mouse model of intranasal carriage of pneumococci to test
whether there is competition between pneumococcal strains.
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Streptococcus pneumoniae colonisation
They found that mice carrying a
serotype 6B strain as resident strain
showed reduced colonisation with a
serotype 23F pneumococcus when
challenged intranasally with the latter
strain. This inhibitory effect could be
overcome by increasing the dose of the
challenge strain.40 Interference in this
complex pattern of interaction and
inhibition by means of vaccination
could have serious and unpredictable
consequences for the composition of
the entire nasopharyngeal population.
Capsule:
polysaccharide
ell wall:
olysaccharide
eichoic acid
holine binding
rotein
Mechanism of colonisation
–
–
–
–
–
IgA1prot
–
– –
–
Pneumococcus
NanA
ChoP
PsaA
CbpA
CbpA
IgA
+
–
+–
+
Mucus layer
GlcNAc
Sialic acid
lacto-Nneotetreose
pIgR
PAFr
Epithelial cell
The pneumococcal outer surface is
Transcytosis
covered by a polysaccharide capsule.
Capsular polysaccharides are highly
heterogeneous, and almost 100
different capsular serotypes have been
described so far.5 The polysaccharide
Cytokine stimulation
capsule is the most important virulence
factor of pneumococci because it Figure 2. Interaction between S pneumoniae and epithelial cells. Neuraminidase (NanA) decreases
protects
the
bacteria
from the viscosity of the mucus and exposes the N-acetyl-glycosamine (GlcNAc) receptors on the
phagocytosis.41 Reduced expression epithelial cells, which can interact with pneumococcal surface-associated proteins such as PsaA. In
response to cytokine stimulation, host epithelial cells upregulate the platelet-activating-factor
results in greater access of antibodies receptors (PAFr). The pneumococcus has increased affinity via its cell-wall phosphocholine (ChoP)
and complement to the pneumococcal for PAFr. Moreover, a second choline-binding protein, CbpA, shows increased affinity for
and
hence
increased immobilised sialic acid and lacto-N-neotreatose, and binds directly to the polymeric Ig receptor
surface,42
clearance by the immune system. (pIgR), which increases migration through the mucosal barrier (transcytosis). Pneumococcal IgA1
Capsular polysaccharides are highly protease cleaves opsonising IgA, which results in a change (neutralisation) of surface charge and
increases the physical proximity of ChoP to the PAFr.
immunogenic. Antibodies against
them protect against infection with the
homologous serotype by induction of opsonophagocytosis. infections.45 This inflammatory cascade changes the type and
The antigenicity of the capsule is type-specific; however, number of receptors on target epithelial and endothelial
cross-reaction can occur because of shared polysaccharides.5 cells. Pneumococcal cell-wall choline shows increased
The layer underneath the capsule, the cell wall, consists affinity for one of these upregulated receptors, the plateletof polysaccharides and teichoic acid and serves as an anchor activating-factor receptor. Binding to this receptor induces
for cell-wall-associated surface proteins. The cell wall is the internalisation of pneumococci and promotes the
cause of the intense inflammatory reaction that accompanies transcellular migration through respiratory epithelium and
pneumococcal infection, since it stimulates the influx of vascular endothelium, resulting in invasion of living bacteria
inflammatory cells and activates the complement cascade (figure 2).46,47 In addition, one of the cell-surface proteins,
and cytokine production.43 The cell wall is believed to be choline-binding protein A (CbpA) shows increased affinity
protected from the host response by the surrounding for immobilised sialic acid and lacto-N-neotetraose on
polysaccharide capsule.
cytokine-activated human cells.48 CbpA directly interacts
Colonisation by S pneumoniae requires adherence to the with the polymeric Ig receptor, which increases migration
epithelial lining of the respiratory tract. Asymptomatic through the mucosal barrier.49 How the pneumococcus
colonisation involves pneumococcal binding to cell-surface escapes endocytosis-mediated killing remains unclear.45,50
carbohydrates (N-acetyl-glycosamine) on non-inflamed The function of IgA1 protease has recently been elucidated
resting epithelium. Adherence to these sugars is mediated by by Weiser and colleagues. They showed increased adherence
cell-wall-associated surface proteins, such as pneumococcal of pneumococci to lung epithelial cells in the presence of
surface adhesin A (PsaA; figure 2). In addition, the surface human IgA. This effect is thought to be brought about by
proteins contribute to the hydrophobic and electrostatic cleavage of opsonising IgA by IgA1 protease, which results in
surface characteristics of pneumococci and might facilitate a change in surface charge and increased physical proximity
adherence to host cells partly through non-specific, of pneumococcal cell-wall choline to the platelet-activatingphysicochemical interactions.44 In general, colonisation is factor receptor.51,52 In addition, CbpA binds to the secretory
not followed by symptomatic disease. Conversion of component of IgA and interacts with the complement
asymptomatic colonisation to invasive disease requires the pathway, thus interfering with the host immune response.49,53
local generation of inflammatory factors such as interleukin Another pneumococcal enzyme, neuraminidase, improves
1 and tumour necrosis factor, as seen in the presence of viral colonisation by cleaving N-acetylneuraminic acid from
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Review
Streptococcus pneumoniae colonisation
mucin, decreasing the viscosity of the mucus.
Neuraminidase also cleaves glycolipids, glycoproteins, and
oligosaccharides, and thus is thought to bring about
exposure of N-acetyl-glycosamine receptors on the host
epithelial cells.54 The neuraminidase activity of viruses such
as influenza and parainfluenza viruses might thereby
contribute to the increased adherence of pneumococci
observed during viral infections.55 Variability in the
composition, expression, or exposure of surface-associated
proteins could explain differences in colonisation and
invasion capacities between strains. The complexity of this
process is underlined by studies in which reversible
phenotypic variation within pneumococcal strains and its
role in host interaction were identified. Transparent phase
variants show greater adherence than opaque variants. This
phenotypic variation is associated with lower expression of
capsule polysaccharides and higher expression of certain
cell-surface proteins and carbohydrate-containing cell-wall
structures.56–58
With increasing knowledge about the mechanisms of
colonisation, surface-associated proteins have become of
major interest as potential vaccine candidates. Although
surface-associated proteins such as pneumolysin and
pneumococcal surface protein A (PspA) elicit protection
against systemic diseases, PsaA and CbpA are promising
candidates for prevention of colonisation.49,59 In theory,
better protection against colonisation and infection with
S pneumoniae might be expected when a combination of
proteins with distinct roles in bacterial virulence is used.
Pneumococcal colonisation in children
Nasopharyngeal colonisation of S pneumoniae in children
mainly depends on age. We investigated the age-dependent
carriage rate in a large cohort of healthy children and
adolescents aged 1–19 years.38 The peak incidence of
pneumococcal colonisation was 55% at the age of 3 years.
There was then a steady decline until a stable prevalence of
8% was observed after the age of 10 years. Although most
other colonisation studies have not extended the age-group
studied into adulthood, those that did have also shown a
decline.60–62 By contrast, the nasopharyngeal niche becomes
colonised during the first year of life. Therefore,
pneumococcal carriage shows an increase before the age of
2 years (table 1).72,81 For example, in a Finnish study the
frequency of nasopharyngeal carriage in children aged
2–24 months increased from 13% for under 6 months to 43%
in children older than 19 months.66 The proportion increased
during respiratory infections to 22–45%, which supports the
theory of greater adherence during (viral) infections.
In the healthy population, risk factors also seem to
determine the frequency of pneumococcal carriage.
Independent determinants for nasopharyngeal colonisation
are ethnicity, crowding, environmental features, and
socioeconomic factors. Socioeconomic and environmental
risk factors include family size (specifically the number of
older siblings), income, smoking (passive and active), and
recent antibiotic use.11,17,28,63,83 Crowding is a major factor in
colonisation and in spread of pneumococcal strains. In
young children, especially, day-care visits are associated with
148
significantly increased colonisation rates (table 1).23,38,74,81–86 In
a study in the Netherlands, the relative risk of
nasopharyngeal colonisation by pneumococci in children
who attended day-care centres compared with children who
were cared for at home was 1·6.23 In addition, that study
showed increased genetic clustering among pneumococcal
isolates, which accords with previous reports.86–88 This
finding supports the hypothesis of increased horizontal
spread of specific pneumococcal strains among attenders at
day-care centres.23 In agreement with these findings,
Raymond and colleagues79 reported a colonisation rate of up
to 82% in infants living in an orphanage. Close relatedness
between the pneumococcal isolates was found in that study,
suggesting frequent horizontal spread.
Ethnic groups at increased risk of pneumococcal
colonisation as well as invasive disease are African American,
native American (Apache and Navajo), and Alaskan native
populations.89 The risk of invasive pneumococcal diseases in
children aged 24–35 months is 64·7 cases per 100 000,
whereas black people in the USA have a rate of 116·4 per 100
000, and native Americans 73–227 cases per 100 000.89 The
risk of invasive disease in the native American population is
increased to such an extent that the US Advisory Committee
for Immunization Practices (ACIP) has recommended
pneumococcal vaccination for this population in all agegroups.1 For children attending day-care centres the risk of
pneumococcal infection is so high that immunisation with a
seven-valent pneumococcal conjugate vaccine (Prevnar,
Wyeth, USA) covering the most prevalent serotypes 4, 6B,
9V, 14, 18C, 19F, and 23F is advised. Pneumococcal
colonisation, especially with antibiotic-resistant bacteria, is
also increased as a result of recent antibiotic treatment.34,83
The selection of antibiotic-resistant pneumococci at the
nasopharynx is commonly assumed to be the cause of the
spread of resistant pneumococcal strains within the
community.77 Consequently, several multidrug-resistant
clones have already spread throughout the world.90,91
Not all risk groups for pneumococcal diseases show
increased rates of colonisation compared with the general
population. For example children with HIV infection and
sickle-cell disease have similar colonisation rates to healthy
children (table 1).70,92 This similarity is a result of the
underlying immune disorder: instead of a defect or
augmented challenge of the primary defence mechanism
against pneumococal invasion, the immune disorder is
related to an impaired response to or clearance mechanism
for pneumococci after invasion has occurred. In children
with HIV/AIDS, the numbers of CD4-positive T cells,
necessary for an appropriate antipolysaccharide response,
are decreased. In children with sickle-cell disease, splenic
function, involved in direct phagocytosis and initiation of
the antipolysaccharide response, is impaired. However, the
primary mucosal barrier, including the mucosal immune
response, is still intact in these patients.92,93
Though variable colonisation rates have been observed
in different areas of the world (table 1), colonisation rates
tend to be higher during respiratory-tract infections and
otitis media and in risk groups such as attenders at day-care
centres. In addition, colonisation rates tend to be higher
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Streptococcus pneumoniae colonisation
when nasopharyngeal samples are obtained via the
oropharynx than by the transnasal approach, though this is
more obvious in healthy children than in those in risk
groups. For future research, we believe the transnasal route
for approaching the nasopharynx is preferable (figure 3).
Serotype distribution among pneumococcal
isolates
The serotype distribution among nasopharyngeal carriage
isolates varies slightly by country, age-group, and type of
cohort. Europe and the US show similar serotype
distributions with minor differences in several serotypes. For
example, in the Netherlands, serotypes 19F (19%), 6B
(16%), 6A (13%), 9V (7%), and 23F (7%) are most
frequently found among children under 3 years of age.23 In
Greece, similarly, the most predominant serotypes among
children younger than 2 years are 6B, 19F, 23F, 14, and
18C;64 and in Finland, serotypes 6B (16%), 23F (14%), 19F
(14%), and 6A (9%) are most prevalent.66 In the USA,
serotypes 6B, 14, 19F, and 23F are also common.94 In Asia,
similar serotypes and serogroups have been found among
nasopharyngeal isolates in healthy children. For example, in
India, the most common serogroups are 6, 14, 19, and 15;63,65
in Vietnam the commonest serogroups are 19, 23, 14, 6, and
18.71 The serogroup distribution in Indonesia is slightly
different, with the most common being 6 (25%) and 23
(21%) followed by 15 (8%), 33 (8%), 19 (6%), 12 (5%), and
3 (4%).67 In Kenya, serotype 13 was with 15, 14, 6B, and 19F
most commonly present.68 In South Africa, a similar
distribution was found with the exception of serotype 13,
which was not found at all.95
No major differences have been found in serotype
distribution between children with risk factors such as
attendance at day-care centres or upper-respiratory-tract
infections and healthy children.23,66,74
By contrast, an important variable is the age-group
investigated. In general, the frequency of vaccine serotypes
declines with age.96 In our study in the Netherlands,38
nasopharyngeal carriage of vaccine-type strains generally
declined from 30% at age 1 year to 3% at 8 years, after which
a stable prevalence was observed until age 19 years. By
contrast, non-conjugate vaccine serotypes, especially
serotypes 3, 8, 10, 11, and 15 showed an increase to the age of
7–10 years, after which there was a delayed decline compared
with the vaccine serotypes.
In general, the serotype distribution among
nasopharyngeal isolates from different parts of the world is
similar. This similarity is also reflected by the potential
conjugate vaccine coverage (table 1). As shown by LloydEvans and colleagues, invasive disease originates from
nasopharyngeal colonisation with the homologous
serotype.97 Therefore, the serotype distribution of
colonisation isolates should be an indicator of invasive
disease, antibiotic resistance profiles, and potential vaccine
coverage. However, certain serotypes and genotypes seem to
cause higher rates of invasive diseases when corrected for
prevalence of nasopharyngeal colonisation.97 Brueggemann
and colleagues98 found serotype-specific and clone-specific
differences in invasive-disease potential with an increased
capacity to cause disease for specific serotype 14 and
18C clones. The most commonly carried serotypes, 6B, 19F,
and 23F, are least invasive, whereas certain non-vaccine
serotypes (8, 38, 33F) are infrequent colonisers but appear to
be more invasive. This is also true for serotypes 5, 7F,
and 1.99,100 This knowledge is extremely important in view of
the replacement of colonising strains observed after
conjugate vaccination. Therefore, surveillance of
pneumococcal invasive disease and colonisation isolates
remains a necessity in those countries where large-scale
pneumococcal vaccination is initiated.
Current vaccine strategies
Figure 3. A nasopharyngeal swab being taken from a 10-year-old girl during
a large cohort study in Rotterdam, Netherlands (September, 2002). The
nasopharynx is approached via the nasal route: the swab is passed gently
back from one nostril along the floor of the nasal cavity until it touches the
posterior wall of the nasopharynx. After gentle rubbing or twisting for 1–2 s,
the swab is withdrawn. The swab is stored in Stuart transport medium and
plated within 6 h onto gentamicin blood agar plates.
THE LANCET Infectious Diseases Vol 4 March 2004
The ACIP has recommended vaccination against
pneumococcal infections for several risk groups. Although
the 23-valent vaccine, with a theoretical coverage of 85–90%
of circulating strains, is immunogenic in adults and children
older than 5 years, young children (<2 years) have a severely
impaired
antibody
response
to
polysaccharide
vaccination.93,101–103 Therefore, the recommendations of the
ACIP in 1997 excluded the major risk group of children
under 2 years of age. The remaining groups were
immunocompetent children older than 2 years at increased
risk of illness and death associated with pneumococcal
disease because of chronic cardiac and pulmonary diseases,
individuals older than 2 years with functional or anatomical
asplenia, and immunocompromised patients older than
2 years.1 Fortunately, the new generation of conjugate
vaccines is highly immunogenic in children under 2 years
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149
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Streptococcus pneumoniae colonisation
Table 2. Conjugate vaccination studies investigating the effect of vaccination on colonisation rate, serotype distribution,
and replacement
Ref Year
Country
Age
Number of Follow-up Risk group Vaccine Vaccination
(months) children
(months)
schedule
Carriage (%)
vaccine types
Carriage (%) of
Replacement
Vaccine Control Vaccine Control
group group group
group
110 2000–01 UK
2
117 1998–99 Netherlands 12–72
607
24–60
..
7-valent 3 x CV + PV
25/43
27/41 10/30
14/32
Not relevant
383
26
Recurrent
7-valent 1–2 x CV + PV
55
55
50
25
Yes
AOM
118 NS
USA
119 1998–99 USA
2
260
10
..
9-valent 3 x CV
41
40
48
60
Yes
7–12
577
11
Native
7-valent 3 x CV
63
65
24
36
Yes
Americans
95
1997
96
1996–97 Israel
South Africa 2
12–35
500
9
..
9-valent 3 x CV
54
61
18
36
Yes
262
24
DCC
9-valent 2 x CV
~65
~70
13
21
Yes
111 (1997)
Israel
2
75
11
..
4-valent 3 x CV + PV†
44–52
52
5–12
30
No
94
1995
USA
2
81
13
..
7-valent‡ 4 x CV
47
53
27
28
Not relevant
112 1994
Israel
263
12
DCC
7-valent 2 x CV
43
57
11
25
No
12–18
CV=conjugate vaccine; NS=not stated; PV=polysaccharide vaccine. *Depending on the season. †Efficacy data do not include the effect of the polysaccharide booster. ‡7-valent
pneumococcal vaccine conjugated to outer membrane protein of N meningitidis.
old. Moreover, these vaccines elicit immunological
memory.104 In several large studies, a seven-valent conjugate
vaccine had almost 100% efficacy against invasive diseases
caused by the included serotypes.105,106 The new vaccines
contain polysaccharides of seven to 11 pneumococcal
serotypes conjugated to a carrier protein inducing a T-celldependent immune response that is present in human
beings from birth. The ACIP has therefore changed the
childhood recommendations for pneumococcal vaccination
in 2000. The current advice is vaccination with the sevenvalent conjugate vaccine Prevnar (Wyeth, USA) for all
children under 2 years of age and in children aged 2–5 years
at increased risk of pneumococcal diseases. In the latter
setting, conjugate vaccination is followed by a
polysaccharide booster, because this step improves
pneumococcal antibody titres in this age-group.107 The
conjugate vaccine is highly effective against invasive diseases
caused by vaccine serotype strains. The efficacy against
mucosal diseases such as pneumonia and otitis media is
much lower and more difficult to measure because cultureproven data are often missing.105,106,108,109 Moreover, several
investigators have shown a significant reduction in
nasopharyngeal carriage of vaccine-type pneumococci as a
result of conjugate vaccination.95,96,110–112 In addition to
individual protection, diminished colonisation is thought to
elicit protection against pneumococcal colonisation and
disease in the vaccinated community—ie, herd immunity.
Dagan and co-workers,113 for example, showed a decreased
colonisation rate in siblings of children attending day-care
centres who were vaccinated with a nine-valent conjugate
vaccine. Moreover, penicillin and multidrug resistance is
common among pneumococcal strains, especially among
the conjugate vaccine serotypes. Therefore, there have been
suggestions that conjugate vaccination will also reduce
resistance among pneumococcal strains in vaccinated
individuals as well as the open community as a result of herd
immunity.89 Recently, Dagan and colleagues114 have shown a
150
significant reduction in penicillin and multidrug resistance
among carriage strains as a result of vaccination with a ninevalent conjugate vaccine.
The vaccines with seven to 11 serotypes inevitably do not
cover all serotypes. Protection also depends on the
geographical area, with potential coverage of the sevenvalent conjugate vaccine for invasive strains of over 85% for
the USA, 60–70% for Europe, and around 55% for Asia,89
although a large proportion of these differences might be
explained by variation in blood-culture practices.115 In
addition to the limited coverage of these conjugate vaccines,
another long-term risk should be considered. Because of the
limited coverage of circulating pneumococcal strains by the
conjugate vaccine, the remaining non-vaccine serotype
strains will actually benefit from this selective
immunological pressure. Replacement may occur, causing a
shift in serotype strains circulating in the population and,
consequently, in disease. Since the start of large-scale
vaccination trials, replacement has been observed in
individuals colonised with pneumococci as well as in
patients with acute otitis media.78,95,109 So far, the effect of this
event on invasive diseases remains unclear. However,
though not yet significant, the first alarming findings have
been reported on partial replacement of invasive strains with
non-vaccine serotypes in vivo.116 In addition, Brueggemann
and co-workers98 have shown a high invasive capacity for
certain non-vaccine serotypes, which may also imply that
replacement of carriage will lead to replacement of disease.
Thus, close monitoring of serotype distribution among
invasive as well as colonisation strains remains of major
importance. Nine studies have investigated the effect of
conjugate vaccination on nasopharyngeal colonisation
(table 2). Two studies found no significant effect on the
overall pneumococcal colonisation nor on vaccine type
carriage.94,110 In the remaining studies, a positive effect of
vaccination was found on colonisation of vaccine-serotype
pneumococci. However, replacement of these strains with
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Streptococcus pneumoniae colonisation
non-vaccine serotypes reduced the effect on overall
pneumococcal colonisation in most cases.
New vaccine strategies
New vaccine strategies focus on the use of pneumococcal
surface-associated proteins. This approach has several
advantages. First, the production of protein vaccines is
expected to be cheap and therefore within reach of
developing countries. Second, a protein-based vaccine is
expected to elicit protection in all age-groups, including
children younger than 2 years. Finally, if highly conserved
proteins or protein epitopes are used as vaccine components,
broad and serotype-independent protection can be expected.
However, the degree and type of protection will be
influenced by the function of the proteins included in the
vaccine. We illustrate this effect by discussing the most
promising protein vaccine candidates.
PspA, one of the family of structurally related cholinebinding surface proteins, can interfere with complement
fixation by blocking recruitment of the alternative pathway
through reduction of the amount of C3b deposited on the
pneumococci, thereby reducing the effectiveness of the
complement-receptor-mediated pathways of clearance.120–123
This process is particularly important when bacterial
invasion has occurred and suggests a significant role for
PspA in the maintenance of invasive pneumococcal disease.
Studies on active immunisation with PspA in animals show a
protective effect against invasive infections and to a lesser
extent against mucosal disease and nasopharyngeal
carriage.124–127 The first phase I vaccination trial with a single
recombinant PspA variant in human beings showed that
broadly cross-reactive antibodies to heterologous PspA
molecules were elicited,128 which were found to protect mice
challenged intraperitoneally with pneumococci.129
Another candidate is PsaA, a member of the family of
metal-binding lipoproteins, part of an ABC transporter
complex thought to be involved in the transport of
manganese into pneumococci.130,131 This protein is mainly
involved in asymptomatic colonisation.45 The first
immunisation studies with PsaA have shown significant
protection against colonisation but limited to modest
protection against invasive infections.59,132–134 Seo and
colleagues135 showed that oral vaccination of mice with PsaA
encapsulated in microalginate microspheres elicited
significant protection against colonisation, pneumonia, and
septicaemia from an oral challenge. These findings suggest
that vaccination with PsaA elicits primary protection against
colonisation with secondary protection against invasive
disease. However, clinical studies on the correlation between
antibodies to PsaA and the risk of pneumococcal acute otitis
media have had contradictory results. Rapola and
co-workers136,137 showed an association between higher titres
of anti-PsaA and lower risk of pneumococcal acute otitis
media, but only in children older than 9 months, whereas in
younger children the risk was increased with higher antiPsaA concentration. These findings suggest a basic difference
among age-groups with respect to protection by antibodies
to PsaA, and perhaps to the origin of the antibody response.
A higher anti-PsaA titre might be associated with increased
THE LANCET Infectious Diseases Vol 4 March 2004
pneumococcal contacts in the past—ie, through
colonisation as well as through infection. Consequently, it
might explain the relation with the underlying increased
susceptibility to pneumococcal acute otitis media rather
then a lower risk of infections.
Pneumolysin is a protein that also contains a cholinebinding domain and is thought to interfere with host
immunity and inflammatory responses by various functions,
including complement fixation and inhibition of phagocyte
function. It also inhibits ciliary activity in the bronchus and
is thus important in pathogenesis of pulmonary infection.138
Knock-out mutagenesis of genes encoding pneumolysin has
suggested a role in virulence, in colonisation as well as in
infection.139–141 Several research groups have described the
protective properties of pneumolysin against challenge with
pneumococci in mice, albeit only against invasive
disease.142,143 The combination of PspA and pneumolysin
yields complementary protection to invasive disease in
animals.59,125 The combination of PsaA and PspA prevents
colonisation and otitis media in animals.125,131 Hence,
depending on the target, differing combinations of vaccine
components can be used. The optimum combination of
proteins to be chosen for vaccination purposes remains to be
investigated.
Alternative routes of vaccination have also been
explored. Several studies124,127,135 have suggested that
administration of a vaccine via the oral or nasal route is as
effective as systemic application. In addition, Lynch and
colleagues144 found that intranasal administration of a
conjugate vaccine plus interleukin 12 not only elicited
protection against invasive disease but also, in contrast to
intramuscular administration, induced protection against
nasal carriage. The latter effect occurs through the induction
of substantial mucosal IgA responses. Mucosal routes of
administration are highly preferable because they are less
invasive and because so many other vaccines are already
administered intramuscularly to children, as part of
community vaccination programmes. Moreover, in contrast
to pneumococcal conjugate vaccines and polysaccharide
vaccines, protection is also expected in children with
HIV/AIDS, even during progression of disease, because of
the intact mucosal immune response in these patients.135
Discussion
Nasopharyngeal colonisation provides an important key to
the burden of pneumococcal disease and its prevention.
Colonisation not only is obligatory for invasive disease, but
also provides the basis for horizontal spread of
pneumococci.
Although the major goal of all vaccine strategies is to
reduce the burden of pneumococcal disease, they involve
also prevention of pneumococcal colonisation. Opinion
about reduction in colonisation ranges from “secondary
aim” to “fortunate side-effect”. However, the importance of
this essential link in pathogenesis has seldom received full
attention.
The natural route of infection with S pneumoniae starts
with colonisation, which may progress to invasive disease
if natural immunological barriers are crossed. Therefore,
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151
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Streptococcus pneumoniae colonisation
a rational aim is to prevent colonisation, thus eliciting
protection against invasive disease. Moreover, prevention of
nasopharyngeal colonisation of S pneumoniae might also
decrease horizontal spread of pneumococcal strains, thus
improving herd immunity.2,113,145 This possibility supports the
use of polysaccharide-based vaccines such as the 23-valent
polysaccharide vaccine and the seven-valent conjugate
vaccine, or future protein-based vaccines consisting of
surface-exposed proteins involved in colonisation and
adherence such as PsaA, CbpA, and neuraminidase.
An alternative to vaccination could be the use of antiattachment agents such as receptor analogues or agents like
xylitol, N-acetylcysteine, or the recently identified
S-carboxymethylcysteine.146 None of these agents results in
complete eradication of pneumococcal colonisation, but the
same is true for vaccination: by prevention of colonisation
without complete eradication of pneumococcal carriage, the
immunological pressure will skew selection of non-covered
serotypes or genotypes. Moreover, if the nasopharyngeal
niche is cleared, replacement with other species might occur.
Veenhoven and colleagues observed that pneumococcal
conjugate vaccination resulted in fewer middle-ear fluid
cultures with vaccine-serotype pneumococci, but in an
increase of three times in cultures positive for S aureus.78
Moreover, we have found competition within the individual
between S aureus and S pneumoniae in healthy children aged
4–9 years.38 Similarly, competition between S pneumoniae
and species such as H influenzae, M catarrhalis, and N
meningitidis has been shown in vitro. A possible solution for
this problem might be to aim strictly for prevention of
invasive disease and leave nasopharyngeal colonisation
unhampered, although mucosal disease can then still occur.
Such disease cannot occur with the currently available
vaccines, but might with future protein-based vaccines
including disease-related proteins such as PspA,
pneumolysin, the phosphate transporter family, and
autolysin.59,147,148 A second option might be to consider the
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